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Cocaine is one of the most abused illicit drugs, and its abuse damages the central nervous system and can even lead directly to death. Therefore, the development of simple, rapid and highly sensitive detection methods is crucial for the prevention and control of drug abuse, traffic accidents and crime. In this work, an electrochemical aptamer-based (EAB) sensor based on the low-temperature enhancement effect was developed for the direct determination of cocaine in bio-samples. The signal gain of the sensor at 10 °C was greatly improved compared to room temperature, owing to the improved affinity between the aptamer and the target. Additionally, the electroactive area of the gold electrode used to fabricate the EAB sensor was increased 20 times by a simple electrochemical roughening method. The porous electrode possesses more efficient electron transfer and better antifouling properties after roughening. These improvements enabled the sensor to achieve rapid detection of cocaine in complex bio-samples. The low detection limits (LOD) of cocaine in undiluted urine, 50% serum and 50% saliva were 70 nM, 30 nM and 10 nM, respectively, which are below the concentration threshold in drugged driving screening. The aptasensor was simple to construct and reusable, which offers potential for drugged driving screening in the real world., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Austria, part of Springer Nature.)

Drug-impaired driving is an increasing public safety concern across Canada, particularly due to the demonstrated increase in use of recreational drugs such as cocaine. Cocaine is a central nervous system stimulant drug; however, it can impair an individual's driving ability in both the stimulant and crash phases. Despite the scientific consensus regarding cocaine's potential for driving impairment, there is relatively little information available regarding blood concentrations and associated observations of impairment in suspected impaired drivers. Retrospective data analysis was performed to evaluate suspected impaired driving cases in which cocaine and/or benzoylecgonine were detected alone, or in combination with other drugs, in blood and urine samples submitted to the Toxicology Section of the Centre of Forensic Sciences with incident dates between 2021 and 2022. Cocaine and/or benzoylecgonine were detected in 46% (blood) and 66% (urine) of the total impaired driving samples submitted. In 41 cases where cocaine and/or benzoylecgonine were the only drug finding in blood, concentrations of cocaine and benzoylecgonine ranged from 0.0073 to 0.26 mg/L (mean 0.096 mg/L) and 0.13 to 5.3 mg/L (mean 2.1 mg/L), respectively. Driving observations reported by the arresting officer in cases where cocaine and/or benzoylecgonine were the only drug finding in blood and urine included the driver being involved in a collision, the vehicle leaving the roadway, erratic driving and the driver being asleep at the wheel; observations of drug impairment reported by the drug recognition expert at the time of driver evaluation included abnormal speech patterns, poor balance/incoordination, abnormal body movements and the individual falling asleep. The results provide concentrations of cocaine and benzoylecgonine observed in suspected impaired drivers, insight into observations that may be associated with prior cocaine use and additional information to inform on the effects of cocaine on driving., (© The Author(s) 2024. Published by Oxford University Press. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site–for further information please contact journals.permissions@oup.com.)

The widespread consumption of cocaine poses a significant threat to modern society. The most effective way to combat this problem is to control the distribution of cocaine, based on its accurate and sensitive detection. Here, we proposed the detection of cocaine in human blood plasma using a combination of surface enhanced Raman spectroscopy and machine learning (SERS-ML). To demonstrate the efficacy of our proposed approach, cocaine was added into blood plasma at various concentrations and drop-deposited onto a specially prepared disposable SERS substrate. SERS substrates were created by deposition of metal nanoclusters on electrospun polymer nanofibers. Subsequently, SERS spectra were measured and as could be expected, the manual distinguishing of cocaine from the spectra proved unfeasible, as its signal was masked by the background signal from blood plasma molecules. To overcome this issue, a database of SERS spectra of cocaine in blood plasma was collected and used for ML training and validation. After training, the reliability of proposed approach was tested on independently prepared samples, with unknown for SERS-ML cocaine presence or absence. As a result, the possibility of rapid determination of cocaine in blood plasma with a probability above 99.5% for cocaine concentrations up to 10 -14 M was confirmed. Therefore, it is evident that the proposed approach has the ability to detect trace amounts of cocaine in bioliquids in an express and simple manner., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The general use of cocaine is increasing in recent years, while the trend for 3,4-methylenedioxymethamphetamine (MDMA) is less clear. The relationship between blood concentrations and impairment is poorly understood, which complicates interpretation. The aims of this study were to report prevalence and blood concentrations of cocaine and MDMA in drugged drivers, and to investigate the relationship between blood concentrations and impairment. Samples of whole blood were collected from apprehended drivers in the period 2000-2022, and a clinical test of impairment (CTI) was simultaneously performed. The samples were initially analyzed for cocaine and MDMA using gas chromatography-mass spectrometry (until 2009 and 2012, respectively), and later using ultra-high-performance liquid chromatography-tandem mass spectrometry. Overall, cocaine was detected in 2,331 cases and MDMA in 2,569 cases. There were 377 and 85 mono cases of cocaine and MDMA, respectively. In the mono cases, the median cocaine concentration was 0.09 mg/L (range: 0.02-1.15 mg/L), and 54% of the drivers were clinically impaired. The median MDMA concentration was 0.19 mg/L (range: 0.04-1.36 mg/L), and 38% were clinically impaired. There was a statistically significant difference in the median cocaine concentration between drivers assessed as not impaired (0.07 mg/L) and drivers assessed as impaired (0.10 mg/L) (P = 0.009). There was also a significant effect of the blood concentration of cocaine (adjusted odds ratio [aOR] = 6.42, 95% confidence interval [CI] = 1.13-36.53, P = 0.036) and driving during the evening/night-time (aOR = 2.17, 95% CI = 1.34-3.51, P = 0.002) on the probability of being assessed as impaired on the CTI. No significant differences were found for MDMA. Many drivers are not assessed as impaired on a CTI following cocaine or especially MDMA use. For cocaine, a relationship between blood concentrations and impairment was demonstrated, but this could not be shown for MDMA., (© The Author(s) 2024. Published by Oxford University Press.)

Cocaine and antidepressants rank high globally in substance consumption, emphasizing their impact on public health. The determination of these compounds and related substances in biological samples is crucial for forensic toxicology. This study focused on developing an innovative analytical method for the determination of cocaine, antidepressants, and their related metabolites in postmortem blood samples, using unmodified commercial Fe 3 O 4 nanoparticles as a sorbent for dispersive magnetic solid-phase extraction (m-d-SPE), coupled with liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) analysis. An aliquot of 100 µL of whole blood and 5 µL of the internal standard pool were added to 30 mg of nanoparticles. The nanoparticles were separated from the sample using a neodymium magnet inserted into a 3D-printed microtube rack. The liquid was then discarded, followed by desorption with 300 µL of 1/1/1 acetonitrile/methanol/ethyl acetate. The sample was vortexed and separated, and 1.5 µL of the organic supernatant was injected into the LC-MS/MS. The method was acceptably validated and successfully applied to 263 postmortem blood samples. All samples evaluated in this study were positive for at least one substance. The most frequent analyte was benzoylecgonine, followed by cocaine and cocaethylene. The most common antidepressants encountered in the analyzed samples were citalopram and fluoxetine, followed by fluoxetine's metabolite norfluoxetine. This study describes the first report of this sorbent in postmortem blood analysis, demonstrating satisfactory results for linearity, precision, accuracy, and selectivity for all compounds. The method's applicability was confirmed, establishing it as an efficient and sustainable alternative to traditional techniques for forensic casework., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH, DE part of Springer Nature.)

Objective: This study examines the results of toxicological tests performed on blood and urine samples collected from suspected drug-impaired drivers in Ontario from 2008 to 2019. The report examines the results of toxicological analysis of the samples submitted, the characteristics of those drivers from whom samples were collected, and the temporal and situational circumstances that led to police investigations and sample collection to better understand drug-impaired driving behavior and to assist in the development and implementation of countermeasure strategies and programs., Methods: Blood and urine samples were sent to the Center of Forensic Sciences where they were analyzed using standardized comprehensive toxicological analysis to test for a wide variety of potentially impairing drugs. Demographic and temporal information for each case from which a sample was collected were also examined to describe the circumstances and characteristics of these driving incidents., Results: During the 12-year period examined, 5,388 samples collected from suspected drug-impaired drivers were analyzed. The number of samples collected increased substantially following the implementation of the Drug Evaluation and Classification Program (DECP) in July 2008, the enactment of legislation facilitating the collection of blood samples from suspects, and the legalization of cannabis for nonmedical purposes in 2018. The number of samples submitted shows temporal correlation with the number of police officers certified as Drug Recognition Experts (DRE) in the province. Over the 12-year period of this study, cannabis was the most frequently detected substance in drivers (52.8% of cases), followed by cocaine (44.3%) and methamphetamine (24.8%). In 80% of cases, more than one substance was detected., Conclusions: Examining the characteristics of suspected drug-impaired drivers, the temporal circumstances, and the drug findings throughout the large geographic area of Ontario and over the extended period of this study enhances our understanding of drug-impaired driving behavior. These characteristics can assist in the development and/or evaluation of enforcement strategies and enhanced countermeasure activities.

Introduction: Cocaine is commonly consumed with ethanol, which leads to the formation of cocaethylene through transesterification. Cocaethylene is an active metabolite of cocaine with a longer duration of action. Literature on the combined toxicity of cocaine, ethanol, and cocaethylene is conflicting. We aimed to compare the acute toxicities of co-exposure to cocaine and ethanol versus cocaine alone in Hong Kong., Methods: This was a retrospective study on acute cocaine toxicities reported to the Hong Kong Poison Control Center from 1 January 2010 to 22 January 2023. Cocaine exposure was confirmed by urine immunoassays/laboratory tests and ethanol co-ingestion was confirmed by blood ethanol concentrations. A serious outcome was defined as a National Poison Data System outcome moderate or above. Univariate analyses and multivariable logistic regression were performed to compare the associations of clinical outcomes with and without ethanol, followed by subgroup analyses of cases with complete data., Results: We analyzed 109 patients (median age 29 years, 71% men, 68% Chinese), of whom 20 had confirmed ethanol co-ingestion (mean blood ethanol concentration 1350 mg/L). Multivariable analysis showed that co-exposure to cocaine and ethanol was associated with a lower risk of serious outcomes (adjusted odds ratio 0.09, 95% confidence interval 0.01-0.77; p = 0.03) after adjusting for age, sex, ethnicity, route of cocaine administration, and physical health status. Subgroup analyses showed similar findings., Conclusions: In contrast to previous studies, we did not identify a higher risk of serious outcomes after co-exposure to cocaine and ethanol compared to cocaine alone in a predominantly Chinese cohort., Competing Interests: Declaration of conflicting interestsThe author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Background: Poisonings resulting from the abuse of drugs currently represent a serious problem for public health. Among the main agents involved, cocaine stands out. It became one of the most abused drugs around the world, and one of the main reasons for visits to the emergency department due to the use of illicit substances. The use of cocaine is primarily in combination with alcoholic beverages. There are few studies that correlate cocaine blood concentration and the severity of clinical manifestations in patients evaluated at Emergency Department. The aim of the present study was to verify the possible relationship between the blood concentration of cocaine and cocaethylene (product of the interaction of cocaine with ethanol) with the severity of the clinical manifestations presented by patients with cocaine intoxication., Methods: Blood levels were measured by high-performance liquid chromatography (HPLC) and the severity of clinical manifestations was assessed using the Stimulant Intoxication Score (SIS). To establish this relationship, Pearson's chi-square statistical test (x 2 ) was used for categorical variables and Student's t for continuous variables, with statistical significance of 5% (p < 0.05)., Results: Of the 81 patients included in the study, 77.8% were men with a mean age of 32.5 years ± 8.5 and mean of SIS 3.4 ± 2.5. Considering the toxicological analysis results, 24.7% of the blood samples were positive. The mean of cocaine and cocaethylene concentrations were 0.34 μg/mL ± 0.45 and 0.38 μg/mL ± 0.34, respectively. The blood concentration of cocaine and cocaethylene has not been shown to be useful information for the treatment and prognosis of patients, but blood levels of these substances at the time of treatment, regardless of their concentration, may be an indicator of severity, showing that any concentrations of these substances should be considered as potentially toxic., Conclusion: The application of the SIS score proved to be an important alternative capable of predicting the severity of the patients due to cocaine intoxication in a fast and simplified way., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Cocaine (COC) is a psychostimulant with a high potential for abuse and addiction. Risk for COC use disorder is driven, in part, by genetic factors. Animal models of addiction-relevant behaviors have proven useful for studying both genetic and nongenetic contributions to drug response. In a previous study, we examined initial locomotor sensitivity to COC in genetically diverse inbred mouse strains. That work highlighted the relevance of pharmacokinetics (PK) in initial locomotor response to COC but was limited by a single dose and two sampling points. The objective of the present study was to characterize the PK and pharmacodynamics of COC and its metabolites (norcocaine and benzoylecgonine) in six inbred mouse strains (I/LnJ, C57BL/6J, FVB/NJ, BTBR T+ tf/J, LG/J and LP/J) that exhibit extreme locomotor responses to cocaine. Mice were administered COC at one of four doses and concentrations of cocaine, norcocaine and benzoylecgonine were analyzed in both plasma and brain tissue at 5 different time points. Initial locomotor sensitivity to COC was used as a pharmacodynamic endpoint. We developed an empirical population PK model that simultaneously characterizes cocaine, norcocaine and benzoylecgonine in plasma and brain tissues. We observed interstrain variability occurring in the brain compartment that may contribute to pharmacodynamic differences among select strains. Our current work paves the way for future studies to explore strain-specific pharmacokinetic differences and identify factors other than PK that are responsible for the diverse behavioral response to COC across these inbred mouse strains., (© 2020 John Wiley & Sons Ltd and International Behavioural and Neural Genetics Society.)

Introduction: Immunoassay (IA) tests are not widely applied in post-mortem samples, since they are based on technologies requiring relatively non-viscous specimens, and compounds originating from the degradation of proteins and lipids during the post-mortem interval can alter the efficiency of the test. However, since the extraction techniques for IA tests are normally rapid and low-cost, IA could be used as near-body drug-screening for the classes of drugs most commonly found in Italy and Europe. In this study, semi-quantitative results on post-mortem whole blood samples obtained through CEDIA analysis (cannabinoids, cocaine, amphetamine compounds, opiates and methadone), were compared with results of confirmatory analysis obtained using GC-MS. Screening cut-offs for all drugs were retrospectively optimized., Methods: Post-mortem whole blood samples from autopsy cases of suspected fatal intoxication were collected over 3 years. Samples were initially analyzed through CEDIA (CEDIA, ILab 650, Werfen). Confirmatory analyses were then performed by GC-MS (QP 2010 Plus, Shimadzu). Screening cut-offs were retrospectively optimized using Receiver Operating Characteristic (ROC) analysis., Results: CEDIA results were available for 125 samples. Two-hundred-eighty-nine (289) positive screening results were found. Among these, 162 positive confirmation results were obtained. Optimized screening cut-offs were as follows: 6.5ng/ml for THC; 4.2ng/ml for THC-COOH; 12.0ng/ml for cocaine; 6.6ng/ml for benzoylecgonine; 6.4ng/ml for opiates; 2.0ng/ml for methadone. Analysis of ROC-curves showed a satisfying degree of separation in all tests except for amphetamine compounds, with areas under the curve (AUC) between 0.915 (THC) and 0.999 (for benzoylecgonine and methadone)., Discussion: The results of the study showed that CEDIA screening at the optimized cut-offs exhibits a very high sensitivity and good specificity and positive predictive value (PPV) for cannabinoids, cocaine and metabolites, opiates and methadone. A high number of false positives (n=19) for amphetamine compounds was observed at the optimized cut-off, resulting in a very low PPV, which is also influenced by the very low number of TP (n=4)., Conclusion: The results of the study show that the CEDIA is a valuable screening test on post-mortem whole blood for cannabinoids, cocaine and metabolites, opiates and methadone, but it is not recommended for amphetamine compounds, due to the high number of false positives. The strengths of the study are the large sample size, the inclusion of post-mortem cases only and the high level of sensitivity and specificity obtained at the optimized cut-offs., Competing Interests: Declaration of Competing Interest The author(s) declare(s) that there is no conflict of interest., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Dried blood spots (DBS) have been considered as complementary matrix in sports drug testing for many years. Especially concerning substances prohibited in-competition only, the added value of DBS collected concomitantly with routine doping control urine samples has been debated, and an increasing potential of DBS has been discussed in the scientific literature. To which extent and under which prerequisites DBS can contribute to enhanced anti-doping efforts is currently evaluated. As a proof-of-principle, two analytical applications, one targeting cocaine/benzoyl ecgonine and the other prednisone/prednisolone, are presented in this perspective to indicate potential added value but also presently existing limitations of the DBS approach., (© 2020 The Authors. Drug Testing and Analysis published by John Wiley & Sons Ltd.)

Liquid chromatography coupled to high-resolution mass spectrometry (HRMS) enables data independent acquisition (DIA) and untargeted screening. However, to avoid the handling of the resulting large dataset, most laboratories in that field still use targeted screening methods, which offer good sensitivity and specificity but are limited to known compounds. The promising field of machine learning offers new possibilities such as artificial neural networks that can be trained to classify large amounts of data. In this proof of concept study, we exemplify such a machine learning approach for raw HRMS-DIA data files. We evaluated a machine learning model using training, validation, and test sets of solvent and whole blood samples containing drugs (of abuse) common in forensic toxicology. For that purpose, different platforms were used. With a feedforward neural network model architecture, a category prediction (blank sample vs. drug containing sample) was aimed for. With the applied machine learning approaches, the sensitivity and specificity, of the validation and test set, for the prediction of sample classes were in a suitable range for an actual use in a (routine) laboratory (e.g. workplace drug testing). In conclusion, this proof of concept study clearly demonstrated the huge potential of machine learning in the analysis of HRMS-DIA data., (© 2020 John Wiley & Sons, Ltd.)

Reliable, feasible analytical methods are needed for forensic and anti-doping testing of cocaine and its most important metabolites, benzoylecgonine, ecgonine methyl ester, and cocaethylene (the active metabolite formed in the presence of ethanol). An innovative workflow is presented here, using minute amounts of dried blood or plasma obtained by volumetric absorptive microsampling (VAMS), followed by miniaturized pretreatment by dispersive pipette extraction (DPX) and LC-MS/MS analysis. After sampling 20 µL of blood or plasma with a VAMS device, the sample was dried, extracted, and loaded onto a DPX tip. The DPX pretreatment lasted less than one minute and after elution with methanol the sample was directly injected into the LC-MS/MS system. The chromatographic analysis was carried out on a C8 column, using a mobile phase containing aqueous formic acid and acetonitrile. Good extraction yield (> 85%), precision (relative standard deviation, RSD < 6.0%) and matrix effect (< 12%) values were obtained. Analyte stability was outstanding (recovery > 85% after 2 months at room temperature). The method was successfully applied to real blood and plasma VAMS, with results in very good agreement with those of fluid samples. The method seems suitable for the monitoring of concomitant cocaine and ethanol use by means of plasma or blood VAMS testing.

We report the on-line coupling of a monolithic molecularly imprinted polymer to nano-liquid chromatography for the selective analysis of cocaine and its main metabolite, benzoylecgonine, in complex biological samples. After the screening of different synthesis conditions, a monolithic molecularly imprinted polymer was in situ synthesized into a 100 μm internal diameter fused-silica capillary using cocaine as template, methacrylic acid as functional monomer, and trimethylolpropane trimethacrylate as cross-linker. Scanning electron microscopy was used to assess the homogeneous morphology of the molecularly imprinted polymer and its permeability was measured. Its selectivity was evaluated by nano-liquid chromatography-ultraviolet, leading to imprinting factors of 3.2 ± 0.5 and 2.2 ± 0.3 for cocaine and benzoylecgonine, respectively, on polymers resulting from three independent syntheses, showing the high selectivity and the repeatability of the synthesis. After optimizing the extraction protocol to promote selectivity, the monolithic molecularly imprinted polymer was successfully on-line coupled with nano-liquid chromatography-ultraviolet for the direct extraction and analysis of cocaine present in spiked human plasma and saliva samples. The repeatability of the obtained extraction recovery, between 85.4 and 98.7% for a plasma sample spiked at 100 ng mL -1 , was high with relative standard deviation values lower than 5.8% for triplicate analyses on each of the three independently synthesized molecularly imprinted polymers. A linear calibration range was achieved between 100 and 2000 ng mL -1  (R 2  = 0.999). Limits of quantification of 14.5 ng mL -1 and 6.1 ng mL -1 were achieved in plasma and urine samples, respectively. The very clean-baseline of the resulting chromatogram illustrated the high selectivity brought by the monolithic molecularly imprinted polymer that allows the removal of a huge peak corresponding to the elution of interfering compounds and the easy determination of the target analyte in these complex biological samples., (Copyright © 2019 Elsevier B.V. All rights reserved.)

The determination of cocaine (COC) and its metabolites ecgonine methyl ester (EME), benzoylecgonine (BZE), norcocaine (NCOC) and cocaethylene (CE) in human plasma is relevant in clinical and forensic toxicology. An efficient extraction and clean-up of plasma specimens for the simultaneous determination of BZE along with COC and basic metabolites is challenging due to their widely different polarities and ionization characteristics. Recently, biocompatible SPME LC tips C18 became commercially available. We applied SPME LC tips C18 to the simultaneous extraction of COC, BZE, EME, NCOC, and CE by direct immersion of the fiber in plasma diluted with a buffer at pH 8.0. Analytes were desorbed from the fiber to methanol containing formic acid and injected into a UPLC-MS/MS system. The assay was linear from 5 to 500 ng mL-1. Precision assays presented CV% in the range of 2.22 to 10.54%, and accuracy was in the range of 93.4-108.1%. The assay requires minimal quantities of plasma and organic solvents, allowing multiple extractions in parallel. Biocompatible SPME is a promising alternative for preparing biological samples prior to drug measurement by UPLC-MS/MS., (© The Author(s) 2019. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Storage and quantitative analysis of small volumes of biofluids are challenging, especially when low concentrations of analytes are to be detected in the presence of complex matrices. In this study, we describe an integrated thread-based approach for stabilizing small blood volumes in the dry-state at room temperature, while also offering direct analysis capabilities via thread spray mass spectrometry. The analytical merits of this novel microsampling platform was demonstrated via the direct analysis of diazepam and cocaine in dried blood samples stored for 42 days. In-situ in-capillary blood processing from hydrophobic threads enabled limits of detection as low as parts-per-quadrillion to be reached. We validated this ultra-sensitivity by analyzing small tissue-like residues collected after pushing a thread through the sample once. The implications of this sample collection, storage, and analysis platform can be extensive with direct applications in forensics and clinical studies., (Published by Elsevier B.V.)

The disposition of drugs and their metabolites have been extensively described in the literature, based primarily on the analysis of plasma and urine. However, there are more limited data on their disposition in whole blood, which is often the only specimen available in forensic investigations and cases of driving under the influence of drugs. In this study, we have, for the first time, established pharmacokinetic properties of cocaine (COC) and its metabolites from concurrently collected whole blood and plasma samples, following a single 100 mg dose of cocaine hydrochloride administered via nasal insufflation to seven healthy volunteers. The median C max of COC and its major metabolites, benzoylecgonine (BZE) and ecgonine methyl ester (EME), were closely related in whole blood and plasma. The median C max for COC in plasma was 379.7 ng/mL (347.5-517.7) and 344.24 ng/mL (271.6-583.2) in whole blood. The median C max for BZE in plasma was 441.2 ng/mL (393.6-475. and 371.18 ng/mL (371.1-477.3) in whole blood, EME was 105.5 ng/mL (93.6-151.8) in plasma and 135.5 ng/mL (87.8-183) in whole blood. Calculated medians of the whole blood to plasma ratio of COC (0.76), BZE (0.98) and EME (1.02) of approximately 1, strongly suggesting that the erythrocyte cell wall presents no barrier to COC and its metabolites. Furthermore, whole blood and plasma concentrations of COC were strongly correlated (R 2  = 0.0914 R = 0.956, p < 0.0001), as was BZE (R 2  = 0.0932 R = 0.965, p < 0.0001) and EME (R 2  = 0.0964R = 0.928, p < 0.0001). The minor oxidative metabolite norcocaine (NCOC) was detected in both whole blood and plasma at concentrations between 1 and 5 ng/mL within 60-180 minutes, suggesting that NCOC could be indicator of recent COC administration. Data from this study have shown for the first time that COC and its metabolites BZE and EME are evenly distributed between plasma and whole blood following controlled single-dose intranasal COC administration., (© 2019 John Wiley & Sons, Ltd.)

Drug screening during pre-transplant evaluations can have major implications for patient care, particularly because drug abuse has been associated with poor transplant outcomes. Although urine drug screening is usually preferred, serum testing is available for situations such as anuria due to end stage renal disease. However, there are few studies evaluating serum drug screening in specific populations such as patients undergoing kidney transplant evaluation. All serum drug screens ordered between January 2015 and November 2017 on patients being evaluated for renal transplant were compared against a large population of serum drug screens ordered from other institutions. Cocaine screening and confirmation results were evaluated to determine false positives. Cocaine screens were positive in 23 of 537 (4.3%) pre-transplant samples, and 211 of 5,115 (4.1%) comparison samples. Confirmation testing demonstrated that 14 (60.9%) pre-transplant samples were false positives, which was significantly (P < 0.01) higher than the rate of false positives in the comparison group (47/211, 22.3%). No common medication or other cross-reacting substance could be identified in the pre-transplant cohort to explain the false-positive results. Although serum cocaine screening had a low overall false-positive rate, the proportion of false positives was significantly higher in pre-transplant patients. Given the poor transplant outcomes associated with drug abuse, failure to properly interpret screening results as being false positives could negatively affect patient care. All members of the transplant team should recognize the importance of confirmation testing in this setting, to avoid unintended consequences due to false-positive screening results., (© The Author(s) 2019. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Objective: Electronic music dance events (EMDEs) at nightclubs attract young adults engaging in high-risk alcohol and other drug (AOD) use. Studies show that most patrons arrive at clubs in groups and that these peer groups influence drinking. Therefore, peer groups are a natural context for preventing risk behaviors. This article examined outcomes of a randomized controlled trial of a group-based mobile intervention at nightclubs, Nightlife Safety Plans (NSP)., Method: The sample comprised 352 groups, consisting of 959 participants (45.3% female) at 41 events across seven nightclubs hosting EMDEs. Club patrons were surveyed anonymously and completed breath tests as they entered and exited clubs. Oral fluid samples collected from patrons at exit assessed drug use. Analyses examining assignment to NSP versus a control condition on fire safety predicted individual- and group-level protective strategy use and AOD use, controlling for background variables., Results: At the individual level, participation in NSP was related to increased protective actions to keep group members safe. No effects were found on actions to keep oneself safe or in response to overuse. At the group level, assignment to NSP was related to a higher average number of group safety strategies. Participation in NSP was associated with lower blood alcohol concentration but unrelated to tetrahydrocannabinol and cocaine., Conclusions: NSP appears to be efficacious for increased protective actions to keep group members safe from overuse and for reduced blood alcohol concentration among EMDE patrons. The findings support the use of an intervention utilizing group-based strategies presented proximal to risk settings.

Brain tissue may serve as a useful supplement to blood in postmortem investigations. However, reference concentrations for central stimulant drugs are scarce in brain tissue. This study involves some frequently used stimulants: amphetamine, cocaine, ephedrine, MDMA and methylphenidate. We present concentrations from brain and blood and brain-blood ratios of the analytes from autopsies. The cases were grouped according to the cause of death: A: The compound solely caused a fatal intoxication. B: The compound contributed to a fatal outcome in combination with other drugs, alcohol or disease. C: The compound was not related to the cause of death. Analyses were carried out using solid-phase extraction and ultra high-performance liquid chromatography. Paired brain and femoral blood concentrations from 133 cases were analysed. Positive correlations were observed for all analytes with correlation coefficients ranging from 0.58 to 0.95. The following median brain-blood ratios were obtained: cocaine 2.0 (range 0.20-7.0), amphetamine 3.2 (range 1.5-4.5), ephedrine 2.3 (range 1.1-6.2), MDMA 3.9 (range 0.92-5.1) and methylphenidate 2.4 (0.92-4.6). The concentrations in femoral blood generally agreed with the literature for all compounds. The metabolite of cocaine, benzoylecgonine, was also quantified in brain and blood from 60 cases, and the median brain-blood ratio was 0.66 with 10-90 percentiles of 0.39-1.27. The results of this study can aid the toxicological investigation in determining the cause of death., (© The Author(s) 2019. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Cocaine is a common illicit stimulant and is mainly metabolized by hydrolysis to benzoylecgonine (BE) and ecgonine methyl ester (EME), but also to minor metabolites like norcocaine, or hydroxy-BE. When ethanol is present, cocaethylene is formed. Dried blood spot (DBS) sampling is a minimally invasive microsampling technique with possible advantages for analyte stability and ease of storage, making it an attractive matrix in forensic and clinical settings. We developed a liquid chromatography-tandem mass spectrometry-based (LC-MS/MS) method for quantifying cocaine, BE, EME, norcocaine, hydroxy-BE, and cocaethylene in DBS. Six-mm punches were extracted with aqueous buffer followed by protein precipitation, evaporation and reconstitution in mobile phase. Separation was achieved on a Polar-RP column (Phenomenex) in a 6-minute gradient including baseline-separation of norcocaine and BE. For MS detection, a QTRAP 5500 (Sciex) was used in positive electrospray ionization (ESI) multiple reaction monitoring (MRM) mode. The method was validated for selectivity, sensitivity [lower limited of quantification (LLOQ) 1.0-5.0 ng/mL], imprecision (≤13.4%, ≤19.6% at LLOQ), accuracy (≤ ± 14.9%), matrix effects, extraction efficiency (≥20.9%), hematocrit effect, volume spotted, punch location, long-term and autosampler stability. Concentrations in DBS from a controlled cocaine administration study in healthy volunteers were compared to whole blood and plasma. Although concentrations correlated moderately to strongly (Spearman's ρ 0.603-0.958), agreement between paired samples was poor, with overestimation of DBS concentrations and wide confidence intervals in Bland-Altman analysis. A possible cause are differences in capillary and venous blood concentrations, with the underlying mechanism requiring further research before DBS analysis for cocaine and its metabolites can be considered equivalent to whole blood or plasma analysis., (© 2018 John Wiley & Sons, Ltd.)

Cocaine (COC) is one of the most widely abused drugs in the world and its sensitive and its reliable measurement in blood is of great importance in the field of forensic and clinical toxicology. Additionally, the determination of COC metabolites such as benzoylecgonine (BZE), cocaethylene (CE), ecgonine methyl ester (EME), and norcocaine (NCOC) are also of complementary diagnostic value. The quantification of COC and metabolites in dried blood spots (DBS) may be an alternative to conventional collection methods with several advantages, including easier, on-site, collection, transportation and storage. In this study, we present a simple and comprehensively validated UPLC-MS/MS assay to measured COC, BZE, EME, NCOC and CE in DBS. The evaluated assay was linear from 5-500 ng mL -1 . Precision assays presented CV% of 1.27-6.82, and accuracy in the range of 97-113.78%. Low haematocrit values had a negative impact in the assay accuracy. COC, BE, NCOC and CE measurements can be made reliably in DBS stored for 14 days at room temperature, as well as at -20 °C and 45 °C. All evaluated compounds can be measured in DBS maintained at -20 °C for 14 days. DBS sampling can be used for the clinical evaluation of the exposure to COC, being an alternative for collection, short-term storage and transportation of blood at room and high temperatures., (Copyright © 2019 Elsevier B.V. All rights reserved.)

The original goal of this study was the employment of surface-enhanced Raman spectroscopy (SERS) for the analysis of real cocaine samples (containing adulterants) on composite Au-TiO 2 nanomaterials to achieve low detection limits suitable for the analysis of illicit drugs and controlled substances and to exploit the photodegradation activity of TiO 2 to recycle the SERS substrate for repeated analyses. The photodegradation (self-cleaning) effects of the Au-TiO 2 composite nanomaterials by ultraviolet (UV) radiation are known. These effects were investigated on large-area SERS substrates immersed in the TiO 2 nanoparticle aqueous suspension. The cocaine samples were measured on electrochemically gold-plated platinum targets. Surprisingly, the intensity of SERS spectra of the pure cocaine did not change after immersion in a suspension of TiO 2 under UV irradiation. However, for some real cocaine samples, the overall intensity of the SERS spectra was even higher after the treatment by TiO 2 and UV radiation as compared to the usual Au substrate. This unexpected signal amplification (valuable for illicit drug detection) was found to be caused mainly by the contained levamisole, which is used as a medical drug and is one of the frequent adulterants of cocaine. Both the sole effect of TiO 2 on the levamisole spectrum intensity and the role of UV irradiation were inspected separately. Finally, an investigation of both the TiO 2 and UV radiation treatments was performed, demonstrating (i) the necessity of both factors for selective SERS signal enhancement of the adulterant and (ii) the revision of general anticipation of the role of TiO 2 in SERS systems.

This article is intended as a brief review or primer about cocaethylene (CE), a pharmacologically active substance formed in the body when a person co-ingests ethanol and cocaine. Reference books widely used in forensic toxicology contain scant information about CE, even though this cocaine metabolite is commonly encountered in routine casework. CE and cocaine are equi-effective at blocking the reuptake of dopamine at receptor sites, thus reinforcing the stimulant effects of the neurotransmitter. In some animal species, the LD50 of CE was lower than for cocaine. CE is also considered more toxic to the heart and liver compared with the parent drug cocaine. The plasma elimination half-life of CE is ~2 h compared with ~1 h for cocaine. The concentrations of CE in blood after drinking alcohol and taking cocaine are difficult to predict and will depend on the timing of administration and the amounts of the two precursor drugs ingested. After an acute single dose of cocaine and ethanol, the concentration-time profile of CE runs on a lower level to that of cocaine, although CE is detectable in blood for several hours longer. A strong case can be made for adding together the concentrations of cocaine and CE in forensic blood samples when toxicological results are interpreted in relation to acute intoxication and the risk of an overdose death., (© The Author(s) 2019. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Background and Rationale: Polydrug use is a widespread phenomenon, especially among adolescents and young adults. Synthetic cathinones are frequently consumed in combination with other drugs of abuse. However, there is very little information regarding the consequences of this specific consumption pattern., Objectives: The aim of this review is to introduce this topic and highlight the gaps in the existing literature. In three different sections, we focus on specific interactions of synthetic cathinones with alcohol, cannabinoids, and the stimulants nicotine and cocaine. We then dedicate a section to the existence of sex and gender differences in the effects of synthetic cathinones and the long-term psychophysiological consequences of adolescent and prenatal exposure to these drugs., Major Findings: Epidemiological studies, case reports, and results obtained in animal models point to the existence of pharmacological and pharmacokinetic interactions between synthetic cathinones and other drugs of abuse. This pattern of polyconsumption can cause the potentiation of negative effects, and the dissociation between objective and subjective effects can increase the combined use of the drugs and the risk of toxicity leading to serious health problems. Certain animal studies indicate a higher vulnerability and effect of cathinones in females. In humans, most of the users are men and case reports show long-term psychotic symptoms after repeated use., Conclusions: The co-use of synthetic cathinones and the other drugs of abuse analyzed indicates potentiation of diverse effects including dependence and addiction, neurotoxicity, and impaired cognition and emotional responses. The motivations for and effects of synthetic cathinone use appear to be influenced by sex/gender. The long-term consequences of their use by adolescents and pregnant women deserve further investigation.

Purpose: The abuse of cocaine and its derivatives presents a likely risk factor for injury. Trauma incurred by cocaine and derivative abusers may be more severe than that incurred by non-users., Objectives: To ascertain the presence of cocaine and its derivatives in trauma patients and to correlate RTS (Revised Trauma Score) and ISS (Injury Severity Score) with the presence of cocaine and its derivatives in blood and urine samples., Methods: All trauma victims treated in an emergency unit between November 11, 2012 and September 15, 2013 were included in the study. Blood and urine samples were collected on admission to hospital. RTS and ISS scores were then compared with the presence or absence of cocaine and its derivatives in the samples. The associations between RTS < 7.84 and ISS > 16 and the independent variables were evaluated by the gross odds ratio values, determined by univariate logistic regression. Multivariate analysis was performed using multivariate logistic regression., Results: Of 453 patients (83.7% male) included in the study, 28.6% presented ISS > 16 and 33.6% presented RTS < 7.84. A total of 435 samples were collected, and 86 (19.8%) provided positive samples for cocaine, 48 (11%) for crack and 69 (15.9%) for cocaethylene. Compared to other patients, drug users showed a greater probability of RTS < 7.84 (2.18 times greater) and a greater probability of ISS > 16 (1.76 times greater)., Conclusion: For the trauma patients included in our study, the use of cocaine and its derivatives was shown to be associated with more severe traumas, as demonstrated by their RTS and ISS scores.

Acetyl fentanyl (N-[1-phenethylpiperidin-4-yl]-N-phenylacetamide) is a potent opioid analgesic with no medicinal uses. We report deaths between 2016 and 2017 at the Medical Examiner's Office in Detroit, MI where acetyl fentanyl was found in the decedent's blood and compare them to previously published deaths between 2015 and 2016. The recent cases (cohort B) had a mean acetyl fentanyl concentration of 0.9 ng/mL (range: 0.1-5.3 ng/mL) and an associated higher concentration of fentanyl along with multiple other drugs present. The older cases (cohort A) had higher concentrations of acetyl fentanyl (mean: 8.9 ng/mL; range: 0.28-37 ng/mL) with lower, yet still toxic, concentrations of fentanyl. We conclude that the cause of death in these recent cases was likely multiple drug toxicity with fentanyl and that the consistently observed lower peripheral blood concentrations of acetyl fentanyl are most likely an artifact in the manufacture of the consumed illicit fentanyl., (© 2018 American Academy of Forensic Sciences.)

Cocaine continues to be one of the most widespread abused illicit drugs in the USA. Rapid methods are needed for the identification and quantitation of cocaine and its metabolites, benzoylecgonine (BE), ecgonine methyl ester (EME) and cocaethylene (CE), in biological specimens by clinical and forensic toxicology laboratories. Presented is a differential ion mobility spectrometry-tandem mass spectrometry (DMS-MS-MS) method for the analysis of cocaine and its major metabolites in human serum that requires minimal sample preparation and no column chromatography. A Shimadzu Nexera X2 ultra-high performance liquid chromatography system was used to infuse the samples into the DMS cell at a rate of 30 μL/min. Separation of cocaine and its metabolites were performed in a SelexION DMS component from Sciex coupled to a QTRAP 6500 with an IonDrive Turbo V source for TurbolonSpray® using acetonitrile as a chemical modifier. Analysis consisted of ramping the CoV from -35 V to -6 V while monitoring the multiple reaction monitoring (MRM) transitions of each analyte. The assay was evaluated for linearity, bias, precision, carryover, interferences and stability. Calibration curves ranged from 10 to 1,000 ng/mL with linear regression correlation coefficients (r2) of 0.9912 or greater for each analyte. The limit of quantitation was set at 10 ng/mL. Intra-day precision (%CV) ranged from 0% to 15% for cocaine, 1% to 19% for BE, 1% to 17% for EME and 0% to 18% for CE. Inter-day precision ranged from 9% to 14% for cocaine, 2% to 17% for BE, 5% to 11% for EME and 5% to 15% for CE. No carryover or interferences were detected. Bland-Altman analysis of previously analyzed specimens by UPLC-MS-MS showed variability of 30% or less. The method demonstrates the applicability of DMS-MS-MS for high throughout analysis of drugs and their metabolites in clinical and forensic toxicology laboratories.

Background: Smuggling of illegal drugs by hiding them inside one's own body, also called body packing, is a worldwide phenomenon. Cocaine is the most frequently transported drug. Body packing is a potentially lethal practice. The most serious complications of body packing are gastrointestinal obstruction or perforation and drug toxicity due to packet leakage or rupture., Case Presentation: A 30-year-old confirmed body packer was brought to our emergency department from jail because of agitation and mydriasis. He presented with a high respiratory rate of 40/min but normal oxygen saturation on ambient air, a heart rate of 116 bpm, a blood pressure of 116/68 mmHg and a temperature of 38.0° Celsius. Blood tests were suggestive of infection, urine analysis was positive for cocaine. Abdominal and thoracic computed tomography scans showed pulmonary infiltrates as a possible focus of infection; signs of bowel obstruction or perforation were absent. Given his clinical presentation, we suspected severe infection rather than massive cocaine intoxication to be the main problem. We therefore withheld immediate surgical decontamination. Instead, we started broad-spectrum antibiotic treatment with piperacillin/tazobactam plus clarithromycin for suspected severe community-acquired pneumonia or abdominal sepsis and treated the patient with intravenous midazolam for symptomatic cocaine intoxication. After detection of urinary pneumococcal antigen, the antibacterial regimen was changed to ceftriaxone and vancomycin for pneumococcal pneumonia. In addition, we found human immunodeficiency virus (HIV) type 1 infection as underlying disease. The patient recovered from his acute illness and was discharged after 7 days of treatment with ceftriaxone plus vancomycin. Antiretroviral therapy was started in an outpatient setting., Conclusions: With this case report, we emphasize the need to look for alternative diagnoses to intoxication and gastrointestinal obstruction in acutely ill body packers with atypical presentation. Special risks, such as underlying HIV infection and potential antimicrobial resistance according to the individual's geographical origin, should be taken into account while treating these patients.

The aims of this study were (1) to identify and quantify cocaine (COC), benzoylecgonine (BE), ecgonine methyl ester (EME), and cocaethylene (CE) in DBS; (2) to compare dried blood spots (DBSs) analytical results with the routine blood analyses; (3) to monitor analytes stability on DBS within a 3-month period. Eighty-five μL of blood from postmortem cases were put on a card for DBS analysis and kept in the dark, at room temperature. Samples were extracted through solid-phase extraction (SPE) cartridges and injected in the liquid chromatography-tandem mass spectrometry (LC-MS/MS) system. The analytical procedure is simple, sensitive, and specific. Limits of detection (LODs) and quantification (LOQs) were calculated at 1.0 and 5.0 ng/mL(g) for COC and CE, and at 0.5 and 2 ng/mL for EME and BE, respectively. Validation parameters fulfilled all the acceptance criteria. Fifty-five postmortem cases were evaluated. Eighteen cases were positive for COC (44-2456 ng/mL) and BE (228-4700 ng/mL), 12 for EME (92-1500 ng/mL), and 11 cases for CE (11-273 ng/mL). Stability was evaluated on 8 cases collected in the period January 2017-January 2018. For each case, 5 DBSs were collected at T0. Four DBSs were analyzed within the 4 following weeks and 1 sample was analyzed after 3 months. The concentrations on DBSs, stored at room temperature, always matched the ones obtained on blood samples kept at -20°C (<20% variation, both at T0 and after 3 months). BE and COC concentrations remained stable after a 3-month storage, EME concentrations slightly increased after 3 weeks in the 2 analyzed samples, while CE provided a less homogeneous stability depending on the sample., (Copyright © 2018 John Wiley & Sons, Ltd.)

It is well-known that 2D dried blood spots on paper offer a facile sample collection, storage, and transportation of blood. However, large volume requirements, possible analyte instability, and difficult sample recovery plague this method, lowering confidence in analyte quantification. For the first time, we demonstrate a new approach using 3D dried blood spheroids for stabilization of small volume blood samples, mitigating these effects without cold storage. Blood spheroids form on hydrophobic paper, preventing interaction between the sample and paper substrate, eliminating all chromatographic effects. Stability of the enzyme alanine transaminase and labile organic compounds such as cocaine and diazepam were also shown to increase in the spheroid by providing a critical radius of insulation. On-surface analysis of the dried blood spheroids using paper spray mass spectrometry resulted in sub-ng/mL limits of detection for all illicit drugs tested, representing 1 order of magnitude improvement compared with analysis from 2D dried blood spots.

Cocaine is a potent stimulant drug widely abused that exists in two forms: as a hydrochloride salt and as a free base (crack). Cocaine and the inactive metabolite benzoylecgonine can be determined to reveal any kind of cocaine use, whereas the pyrolysis product anhydroecgonine methyl ester (AEME) can be determined to reveal crack smoking. There are many bioanalytical LC-MS/MS methods used for the determination of cocaine, metabolites and AEME. In these methods, chromatographic separation is usually performed by HPLC and sample preparation by solid phase extraction. For the first time, an UHPLC-MS/MS method for the simultaneous determination of cocaine, benzoylecgonine, cocaethylene and AEME in blood using a sample preparation by liquid-liquid extraction was developed and validated. Extraction recoveries were approximately 80%, 40%, 80% and 80%, respectively, obtained by using a mixture of MTBE/2-propanol (70:30, v:v). Chromatographic separation was performed on a core shell biphenyl UHPLC column (100×2.1mm ID, 1.7μm particles). Method validation showed that the method is precise, accurate, robust and sensitive for its purposes. Limit of quantification (LOQ) concentrations were 0.7-1.5ng/mL. The method was used to determine cocaine, benzoylecgonine, cocaethylene and AEME in 22 blood samples collected from victims of sudden, unexpected or violent death in Sao Paulo (Brazil). Concentrations ≥LOQ were observed in 19, 21, 10 and 10 of these samples, respectively., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Adulteration is a common practice in the illicit drugs market, but the psychoactive and toxic effects provided by adulterants are clinically underestimated. Coca-paste (CP) is a smokable form of cocaine which has an extremely high abuse liability. CP seized samples are sold adulterated; however, qualitative and quantitative data of CP adulteration in forensic literature is still scarce. Besides, it is unknown if adulterants remain stable when CP is heated. This study was designed to report the chemical content of an extensive series of CP seized samples and to demonstrate the stability (i.e., chemical integrity) of the adulterants heated. To achieve this goal, the following strategies were applied: (1) a CP adulterated sample was heated and its fume was chemically analyzed; (2) the vapor of isolated adulterants were analyzed after heating; (3) plasma levels of animals exposed to CP and adulterants were measured. Ninety percent of CP seized samples were adulterated. Adulteration was dominated by phenacetin and caffeine and much less by other compounds (i.e., aminopyrine, levamisole, benzocaine). In the majority of CP analyzed samples, both cocaine and caffeine content was 30%, phenacetin 20% and the combination of these three components reached 90%. Typical cocaine pyrolysis compounds (i.e., BA, CMCHTs, and AEME) were observed in the volatilized cocaine and CP sample but no pyrolysis compounds were found after isolated adulterants heating. Cocaine, phenacetin, and caffeine were detected in plasma. We provide current forensic data about CP seized samples and demonstrated the chemical integrity of their adulterants heated.

Background: TV-1380 is a rationally mutated, human BChE fused to human serum albumin that has high hydrolytic enzymatic activity against cocaine and as well as an extended elimination half-life., Objective: The present studies examined the safety of TV-1380 and its protective effect when given to monkeys alone or concomitantly with cocaine and ethanol., Methods: A set of studies was conducted in monkeys with TV-1380. The parameters tested included telemetric assessment of cardiovascular parameters, clinical pathology, plasma analysis of cardiac troponin I, ex-vivo analyses of cocaethylene and PK analysis of serum concentrations of TV-1380, cocaine and its metabolites, and histopathological examinations., Results: TV-1380 treatment in monkeys was well tolerated. TV-1380 pretreatment prior to cocaine significantly attenuated the cardiac effects of cocaine and reduced cocaine-induced elevations in serum cardiac troponin I. TV-1380 changed the metabolic fate of cocaine resulting in decreased exposure to benzoylecgonine, while increasing the exposure to ecgonine methyl ester in plasma.TV-1380 reduced the plasma levels of the toxic metabolite cocaethylene formed after co-administration of ethanol and cocaine., Conclusion: The results of this study demonstrate that TV-1380 not only accelerates the elimination of cocaine, but also protects the treated animal from the cardiac effects of cocaine, and inhibits the formation of the toxic cocaethylene metabolite when cocaine is given together with ethanol, supporting further clinical development of modified BChE products as possible treatments for cocaine abuse., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Cocaine is one of the most used illegal recreational drugs. Developing an on-site test for cocaine use detection has been a focus of research effort, since it is essential to the control and legal action against drug abuse. Currently most of cocaine detection methods are time-consuming and require special or expensive equipment, and the detection often suffers from high cross-reactivity with cocaine metabolites and relative low sensitivity with the best limit of detection reported at sub nanomolar (nM) level. In this work, an aptasensor has been developed using capacitive monitoring of sensor surface incorporating alternating current electrokinetics effects to speed up molecular transport and minimize matrix effects. The aptasensor is rapid, low cost, highly sensitive and specific as well as simple-to-use for the detection of cocaine from serum. The assay has a sample-to-result time of 30 s, a limit of detection of 7.8 fM, and a linear response for cocaine ranging from 14.5fM to 14.5pM in standard buffer, which are great improvements from other reported cocaine sensors. Special buffer is used for serum cocaine detection, and a limit of detection of 13.4 fM is experimentally demonstrated for cocaine spiked in human serum (equivalent to 1.34pM cocaine in neat serum). The specificity of the biosensor is also demonstrated with structurally similar chemicals, ecgonine ethyl ester and methylecgonidine. This biosensor shows high promise in detection of low levels of cocaine from complex matrices., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Our laboratory received segments of umbilical cord that originated from identical twins for routine toxicology analysis. The specimens were analyzed multiple times by liquid chromatography tandem mass spectrometry. The umbilical cord from newborn #1 was positive for hydromorphone only (1.06 ng/g), and the umbilical cord from newborn #2 was positive for hydromorphone (0.81 ng/g) and benzoylecgonine (5.41 ng/g). The hydromorphone results are consistent with maternal administration of hydromorphone; however, the cause of the discrepant benzoylecgonine results in the umbilical cords from the identical twins is unknown.

The pharmacokinetic profile of oral cocaine has not been fully characterized and prospective data on oral bioavailability are limited. A within-subject study was performed to characterize the bioavailability and pharmacokinetics of oral cocaine. Fourteen healthy inpatient participants (six males) with current histories of cocaine use were administered two oral doses (100 and 200 mg) and one intravenous (IV) dose (40 mg) of cocaine during three separate dosing sessions. Plasma samples were collected for up to 24 h after dosing and analyzed for cocaine and metabolites by gas chromatography-mass spectrometry. Pharmacokinetic parameters were calculated by non-compartmental analysis, and a two-factor model was used to assess for dose and sex differences. The mean ± SEM oral cocaine bioavailability was 0.32 ± 0.04 after 100 and 0.45 ± 0.06 after 200 mg oral cocaine. Volume of distribution (Vd) and clearance (CL) were both greatest after 100 mg oral (Vd = 4.2 L/kg; CL = 116.2 mL/[min kg]) compared to 200 mg oral (Vd = 2.9 L/kg; CL = 87.5 mL/[min kg]) and 40 mg IV (Vd = 1.3 L/kg; CL = 32.7 mL/[min kg]). Oral cocaine area-under-thecurve (AUC) and peak concentration increased in a dose-related manner. AUC metabolite-to-parent ratios of benzoylecgonine and ecgonine methyl ester were significantly higher after oral compared to IV administration and highest after the lower oral dose. In addition, minor metabolites were detected in higher concentrations after oral compared to IV cocaine. Oral cocaine produced a pharmacokinetic profile different from IV cocaine, which appears as a rightward and downward shift in the concentration-time profile. Cocaine bioavailability values were similar to previous estimates. Oral cocaine also produced a unique metabolic profile, with greater concentrations of major and minor metabolites.

Reliable field testing devices for psychoactive drugs would be useful tools for the police for detecting drug-impaired drivers. The Norwegian Mobile Police Service (NMPS) started using Dräger DrugTest 5000 (DDT5000) in 2015 as an on-site screening instrument for drugs in samples of oral fluid. The aim of this study was to compare the results of field testing of DDT5000 with drug findings in blood and oral fluid samples taken from drivers suspected for driving under the influence of drugs (DUID). In total, 369 drivers were included in this field testing; blood samples were obtained from all of them, while oral fluid samples were collected with the Intercept device from 301 of them. The median time from field testing with DDT5000 and collection of blood and oral fluid samples was 50 min. The proportions of false positive results with DDT5000 compared to findings in blood samples above the Norwegian legal per se limits were for cannabis 14.5%, amphetamine 23.2%, methamphetamine 38.4%, cocaine 87.1%, opiates 65.9% and benzodiazepines 36.4%. The proportions of false negatives were for cannabis 13.4%, amphetamine 4.9%, methamphetamine 6.1%, cocaine 0.0%, opiates 0.0% and benzodiazepines 18.8%. Among drivers who had drug concentrations above the legal limits in blood, the proportion who tested positive using DDT5000 was 82.9% for THC, 90.8% for amphetamine, 75.7% for methamphetamine, 100.0% for cocaine, 100.0% for opiates and 37.2% for benzodiazepines. In cases with false-positive DDT5000 results compared to blood, traces of drugs were most often found in oral fluid. The DDT5000 did not absolutely correctly identify DUID offenders due to fairly large proportions of false-positive or false-negative results compared to drug concentrations in blood. The police reported that DDT5000 was still a valuable tool in identifying possible DUID offenders, resulting in more than doubling the number of apprehended DUID offenders.

A molecularly imprinted polymer - Mn-doped ZnS quantum dot-based fluorescence probe for cocaine abuse screening has been prepared and applied to complex samples such as serum and oral fluid. The fluorescent sensing material was prepared by anchoring a selective MIP for COC on the surface of polyethylene glycol (PEG) modified Mn-doped ZnS quantum dots (QDs). Simple and low cost methods have thus been optimized for assessing cocaine abuse in serum and oral fluid by monitoring fluorescence quenching when cocaine (COC) is present (optimized operating conditions with 1.5mL of 200mgL -1 MIP-coated QDs solution, pH 5.5, and 15min before fluorescence scanning). The matrix effect was found to be important when analyzing oral fluid and serum, and several strategies based on centrifugation for oral fluid and solid phase extraction (SPE) for serum were explored. Two analytical methods were developed for oral fluid. The first one (direct method) requires a centrifugation step (6°C, 4000rpm, 20min) to avoid the matrix effect, and allows for cocaine determination by using an aqueous calibration (1:20 dilution). The second method was developed for oral fluid sampled by Salivette devices, and also requires a further centrifugation (6°C, 4000rpm, 20min) of the recovered oral fluid. This method, however, requires the standard addition technique (1:20 dilution) because of the existence of the matrix effect. Regarding serum samples, a direct method (serum dilution) was not possible, and an SPE procedure was needed to avoid the matrix effect (use of aqueous calibration). The limits of detection and quantification when using the Salivette method were 0.035mgL -1 and 0.117mgL -1, respectively; whereas, 0.015mgL -1 (LOD) and 0.050mgL -1 (LOQ) were obtained for serum., (Copyright © 2018 Elsevier B.V. All rights reserved.)

The use of oral fluid (OF) as an alternative specimen for drug analysis has become very popular in forensic toxicology. Many clinical studies have evaluated the correlations between concentrations of cocaine and its metabolites in OF and other matrices, but results have shown high variability. In addition, there are no data available regarding the correlations between biomarkers of crack-cocaine use in different matrices. This study evaluated the relationship between concentrations of cocaine/crack-cocaine biomarkers in OF, urine and plasma samples collected from cocaine users. All samples were analyzed for the presence of cocaine (COC), benzoylecgonine (BZE) and anhydroecgonine (AEC) by a validated liquid chromatography-mass spectrometry method. Median COC, BZE and AEC concentrations ranged from 4.20 to 33.26 ng/mL, from 13.03 to 3,615.86 ng/mL and from 7.40 to 1,892.5 ng/mL across matrices, respectively. The relationship between drug concentrations in OF versus plasma (OF/P) and OF versus urine (OF/U) was evaluated by their coefficients of determination (R2). Least-squares regression analyses demonstrated significant correlations between OF/P and OF/U for cocaine and BE (P < 0.05), with R2 = 0.17, 0.07 for cocaine and R2 = 0.73, 0.45 for BE, respectively. The correlation coefficients (r) found for BZE, COC and AEC in OF/P and OF/U were 0.85 and 0.67 (P < 0.05); 0.41 and 0.26 (P < 0.05); and 0.30 and -0.37 (P > 0.05), respectively. Many factors contribute to the variability of drug correlation ratios in studies involving random samples, including uncertainty about the time of last administration and dosage. Overall, we found significant R2 values for COC and BZE in OF/P and OF/U, but not for AEC. Despite the good correlations found in some cases, especially for BZE, the large variation in drug concentrations seen in this work suggests that OF concentrations should not be used to estimate concentrations of COC, BZE or AEC in plasma and/or urine.

Here we present a potentiometric sensor for cocaine detection based on molecularly imprinted polymer nanoparticles (nanoMIPs) produced by the solid-phase imprinting method. The composition of polymers with high affinity for cocaine was optimised using molecular modelling. Four compositions were selected and polymers prepared using two protocols: chemical polymerisation in water and UV-initiated polymerisation in organic solvent. All synthesised nanoparticles had very good affinity to cocaine with dissociation constants between 0.6nM and 5.3nM. Imprinted polymers produced in organic solvent using acrylamide as a functional monomer demonstrated the highest yield and affinity, and so were selected for further sensor development. For this, nanoparticles were incorporated within a PVC matrix which was then used to prepare an ion-selective membrane integrated with a potentiometric transducer. It was demonstrated that the sensor was able to quantify cocaine in blood serum samples in the range of concentrations between 1nM and 1mM., (Copyright © 2017 Elsevier B.V. All rights reserved.)

A detailed description is given of the development and validation of a fully automated in-line solid-phase extraction-liquid chromatography-tandem mass spectrometry (SPE-LC-MS/MS) method capable of detecting 90 central-stimulating new psychoactive substances (NPS) and 5 conventional amphetamine-type stimulants (amphetamine, 3,4-methylenedioxy-methamphetamine (MDMA), 3,4-methylenedioxy-amphetamine (MDA), 3,4-methylenedioxy-N-ethyl-amphetamine (MDEA), methamphetamine) in serum. The aim was to apply the validated method to forensic samples. The preparation of 150μL of serum was performed by an Instrument Top Sample Preparation (ITSP)-SPE with mixed mode cation exchanger cartridges. The extracts were directly injected into an LC-MS/MS system, using a biphenyl column and gradient elution with 2mM ammonium formate/0.1% formic acid and acetonitrile/0.1% formic acid as mobile phases. The chromatographic run time amounts to 9.3min (including re-equilibration). The total cycle time is 11min, due to the interlacing between sample preparation and analysis. The method was fully validated using 69 NPS and five conventional amphetamine-type stimulants, according to the guidelines of the Society of Toxicological and Forensic Chemistry (GTFCh). The guidelines were fully achieved for 62 analytes (with a limit of detection (LOD) between 0.2 and 4μg/L), whilst full validation was not feasible for the remaining 12 analytes. For the fully validated analytes, the method achieved linearity in the 5μg/L (lower limit of quantification, LLOQ) to 250μg/L range (coefficients of determination>0.99). Recoveries for 69 of these compounds were greater than 50%, with relative standard deviations≤15%. The validated method was then tested for its capability in detecting a further 21 NPS, thus totalling 95 tested substances. An LOD between 0.4 and 1.6μg/L was obtained for these 21 additional qualitatively-measured substances. The method was subsequently successfully applied to 28 specimens from routine forensic case work, of which 7 samples were determined to be positive for NPS consumption., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Illicit cocaine seizures are often compared to each other by using gas chromatography-mass spectrometry (GC-MS) data from cocaine alkaloid compounds to determine whether two specimens originate from the same production batch or not. This can provide intelligence or investigative information at the early stages of an investigation or evidence in court. Traditional classification methods assume high stability of all alkaloids, use all of them to calculate the correlation between two profiles and use a threshold to classify samples. Unstable alkaloids will have a strong influence on the performance. We show that comparing each alkaloid target compound individually improves the classification. Unfortunately, it requires normalization and is also sensitive to the stability. Instead we suggest to use ratios of all possible pairwise combinations of the GC-MS peaks. These ratios are scale free and directly comparable between samples. The peaks can be given different weights in the comparison of profiles using appropriate classification methods and we show that randomForest classification using these ratios have a high and reproducible performance in comparison with other methods. The performance of this method is not affected by noise, transformation or normalization and should be considered for future comparison of chromatographic profiles in general.

Background: Cocaine is a well-known risk factor for acute cardiac events, but the effects in users outside of acute events are less clear. We investigated a possible association between cocaine use and the concentration of a novel biomarker for cardiac stress and heart failure, ST2., Methods: A case-control study was conducted to compare ST2 concentrations by the presence of cocaine in patients presenting for care, but not cardiac care, at an urban safety net hospital., Results: In samples taken from 100 cocaine-positive and 100 cocaine-negative patients, the presence of cocaine was associated with ST2 concentrations>35ng/mL. Serum concentrations of benzoylecgonine, a major cocaine metabolite, were significantly correlated with ST2 concentrations., Conclusions: Cocaine use is associated with subclinical cardiac stress and damage outside of acute cardiac events. This information could add to better stratification of cocaine users with elevated ST2 concentrations who may be at higher risk for developing heart failure and other cardiac complications., (Copyright © 2017 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.)

The need to calibrate to correct for sensor-to-sensor fabrication variation and sensor drift has proven a significant hurdle in the widespread use of biosensors. To maintain clinically relevant (±20% for this application) accuracy, for example, commercial continuous glucose monitors require recalibration several times a day, decreasing convenience and increasing the chance of user errors. Here, however, we demonstrate a "dual-frequency" approach for achieving the calibration-free operation of electrochemical biosensors that generate an output by using square-wave voltammetry to monitor binding-induced changes in electron transfer kinetics. Specifically, we use the square-wave frequency dependence of their response to produce a ratiometric signal, the ratio of peak currents collected at responsive and non- (or low) responsive square-wave frequencies, which is largely insensitive to drift and sensor-to-sensor fabrication variations. Using electrochemical aptamer-based (E-AB) biosensors as our test bed, we demonstrate the accurate and precise operation of sensors against multiple drugs, achieving accuracy in the measurement of their targets of within better than 20% across dynamic ranges of up to 2 orders of magnitude without the need to calibrate each individual sensor.

A universal aptameric system based on the taking advantage of double-stranded DNA/perylene diimide (dsDNA/PDI) as the signal probe was developed for multiplexed detection of small molecules. Aptamers are single-stranded DNA or RNA oligonucleotides which are selected in vitro by a process known as systematic evolution of ligands by exponential enrichment. In this work, we synthesized a new kind of PDI and reported this aggregated PDI could quench the double-stranded DNA (dsDNA)-labeled fluorophores with a high quenching efficiency. The quenching efficiencies on the fluorescence of FAM, TAMRA and Cy5 could reach to 98.3%±0.9%, 97.2%±0.6% and 98.1%±1.1%, respectively. This broad-spectrum quencher was then adopted to construct a multicolor biosensor via a label-free approach. A structure-switching-triggered enzymatic recycling amplification was employed for signal amplification. High quenching efficiency combined with autocatalytic target recycling amplification afforded the biosensor with high sensitivity towards small analytes. For other targets, changing the corresponding aptamer can achieve the goal. The quencher did not interfere with the catalytic activity of nuclease. The biosensor could be manipulated with similar sensitivity no matter in pre-addition or post-addition manner. Moreover, simultaneous and multiplexed analysis of several small molecules in homogeneous solution was achieved, demonstrating its potential application in the rapid screening of multiple biotargets., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Development of simple, sensitive, and rapid method for cocaine detection is important in medicine and drug abuse monitoring. Taking advantage of fluorescence anisotropy and aptamer, this study reports a direct fluorescence anisotropy (FA) assay for cocaine by employing an aptamer probe with tetramethylrhodamine (TMR) labeled on a specific position. The binding of cocaine and the aptamer causes a structure change of the TMR-labeled aptamer, leading to changes of the interaction between labeled TMR and adjacent G bases in aptamer sequence, so FA of TMR varies with increasing of cocaine. After screening different labeling positions of the aptamer, including thymine (T) bases and terminals of the aptamer, we obtained a favorable aptamer probe with TMR labeled on the 25th base T in the sequence, which exhibited sensitive and significant FA-decreasing responses upon cocaine. Under optimized assay conditions, this TMR-labeled aptamer allowed for direct FA detection of cocaine as low as 5 μM. The maximum FA change reached about 0.086. This FA method also enabled the detection of cocaine spiked in diluted serum and urine samples, showing potential for applications. Graphical Abstract The binding of cocaine to the TMR-labeled aptamer causes conformation change and alteration of the intramolecular interaction between TMR and bases of aptamer, leading to variance of fluorescence anisotropy (FA) of TMR, so direct FA analyis of cocaine is achieved.

A sensitive high-performance liquid chromatography (HPLC) assay with dual UV detection has been developed and validated for the simultaneous quantification of methadone and cocaine in rat serum and brain tissue samples. Liquid-liquid extraction using hexanes was applied for samples extraction with Levo-Tetrahydropalmatine (L-THP) as the internal standard. Chromatographic separation of the analytes was achieved on a reversed-phase Waters Symmetry ® C 18 column (150mm×4.6mm, 5μm). A gradient elution was employed with a mobile phase consisting of 5mM potassium phosphate containing 0.1% triethylamine (pH=6.5) (A) and acetonitrile (B) with a flow rate of 1mL/min. UV detection was employed at 215nm and 235nm for the determination of methadone and cocaine, respectively. The calibration curves were linear over the range of 0.05-10μg/mL for both methadone and cocaine. The assay was validated according to FDA guidelines for bioanalytical method validation and results were satisfactory and met FDA criteria. Inter-day accuracy values of serum and brain samples ranged from 96.97 to 105.59% while intra-day accuracy values ranged from 91.49 to 111.92%. Stability assays showed that both methadone and cocaine were stable during sample storage, preparation, and analytical procedures. The method was successfully used to analyze biological samples obtained from a drug- drug interaction pharmacokinetics (PK) study conducted in rats to investigate the effect of methadone on cocaine PK. Our method not only can be used for bioanalysis of samples obtained from rats but also can potentially be applied to human biological serum samples to monitor compliance to methadone maintenance therapy (MMT) and to detect possible cocaine-methadone co-abuse., (Copyright © 2017 Elsevier B.V. All rights reserved.)